First report of target-site resistance to ACCase-inhibiting herbicides in Bromus tectorum L.

BACKGROUND: The prevalent and repeated use of acetyl-coenzyme A carboxylase (ACCase)-inhibiting herbicides for Bromus tectorum L. control in fine fescue (Festuca L. spp) grown for seed has selected ACCase-resistant B. tectorum populations. The objectives of this study were to (1) evaluate the response of nine B. tectorum populations to the ACCase inhibitors clethodim, sethoxydim, fluazifop-P-butyl, and quizalofop-P-ethyl and the acetolactate synthase (ALS) inhibitor sulfosulfuron and (2) characterize the resistance mechanisms. RESULTS: Bromus tectorum populations were confirmed to be resistant to the ACCase-inhibiting herbicides tested. The levels of resistance varied among the populations for clethodim (resistance ratio, RR = 5.1-14.5), sethoxydim (RR = 18.7-44.7), fluazifop-P-butyl (RR = 3.1-40.3), and quizalofop-P-ethyl (RR = 14.5-36). Molecular investigations revealed that the mutations Ile2041Thr and Gly2096Ala were the molecular basis of resistance to the ACCase-inhibiting herbicides. The Gly2096Ala mutation resulted in cross-resistance to the aryloxyphenoxypropionate (APP) herbicides fluazifop-P-butyl and quizalofop-P-ethyl, and the cyclohexanedione (CHD) herbicides clethodim, and sethoxydim, whereas Ile2041Thr mutation resulted in resistance only to the two APP herbicides. All B. tectorum populations were susceptible to sulfosulfuron (RR = 0.3-1.7). CONCLUSIONS: This is the first report of target-site mutations conferring resistance to ACCase-inhibiting herbicides in B. tectorum. The results of this study suggest multiple evolutionary origins of resistance and contribute to understanding the patterns of cross-resistance to ACCase inhibitors associated with different mutations in B. tectorum. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Molecular and Alkaloid Characterization of Claviceps purpurea Sensu Lato From Grass Seed Production Areas of the U.S. Pacific Northwest.

Ergot, caused by Claviceps purpurea sensu lato, is an economically important seed replacement disease of Kentucky bluegrass (Poa pratensis) and perennial ryegrass (Lolium perenne) seed crops. C. purpurea sensu stricto is considered the primary Claviceps species responsible, but genetic diversity and cryptic species within C. purpurea sensu lato have previously been reported. Fifty-six C. purpurea sensu lato isolates collected from P. pratensis (n = 21) and L. perenne (n = 35) in Oregon and Washington between 2010 and 2014 were characterized via random amplified polymorphic DNA (RAPD), partial internal transcribed spacer (ITS), ß-tubulin and elongation factor-1α (EF-1α) sequences, conidial size, and ergot alkaloid chemotype. Based on RAPD analysis, seven isolates from P. pratensis and 33 isolates from L. perenne collected in Oregon corresponded to C. purpurea sensu stricto, and 13 isolates collected from P. pratensis in Washington and Oregon were identified as C. humidiphila. Partial ITS, ß-tubulin, and EF-1α sequences identified 10 isolates from P. pratensis as C. humidiphila, and seven isolates from P. pratensis and 33 isolates from L. perenne were identified as C. purpurea sensu stricto. Several isolates generated ambiguous RAPD bands or sequences that prevented identification. Ergot alkaloid chemotype profiling found that ergocornine and its epimer were predominant in sclerotia from P. pratensis, whereas ergotamine and its epimer were most abundant in sclerotia from L. perenne. This study confirms the presence of the C. purpurea sensu lato species complex in the U.S. Pacific Northwest and suggests that more research is needed to characterize and mitigate Claviceps spp. infection of grass seed crops in North America.

Detection and Quantification of Airborne Claviceps purpurea sensu lato Ascospores from Hirst-Type Spore Traps using Real-Time Quantitative PCR.

The U.S. Pacific Northwest states of Oregon and Washington are major producers of cool-season grass seed. Ergot, caused by fungi in the Claviceps purpurea sensu lato group, is an important seed replacement disease of grass worldwide. Microscopic methods that are currently used to quantify airborne Claviceps ascospores captured by spore traps are not currently rapid enough to allow for detecting and reporting of spore numbers in a timely manner, hindering growers from using this information to help manage ergot. We developed a SYBR Green real-time quantitative polymerase chain reaction (qPCR)-based assay for fast and efficient detection and quantification of C. purpurea sensu lato ascospores from Hirst-type spore traps. Species-specificity of the qPCR assay was confirmed against 41 C. purpurea sensu lato isolates collected from six hosts and six other Claviceps spp. Significant relationships were observed between cycle threshold (Ct) values and standard curves of serial dilutions of DNA ranging from 1 pg to 10 ng (R2 = -0.99; P = 0.0002) and DNA extracted from a conidial suspension representing 8 to 80,000 conidia (R2 = -0.99; P = 0.0004). Ct values from qPCR were significantly correlated with results from microscopic examination of spore trap samples from the field (r = -0.68; P < 0.0001) and the procedure was able to detect a single ascospore from spore trap tape samples. The qPCR procedure developed in this study provided a means for quantifying airborne Claviceps ascospores that was highly specific and useful over a wide range of spore densities, and could be performed in a matter of hours instead of days. The qPCR assay developed in this study could be part of an integrated pest management approach to help grass seed growers make risk-based fungicide application decisions for ergot management in grass grown for seed.

Spatial and Temporal Dynamics of Aphids (Hemiptera: Aphididae) in the Columbia Basin and Northeastern Oregon.

Aphid species, such as the potato aphid, Macrosiphum euphorbiae Thomas, and the green peach aphid, Myzus persicae Sulzer, are routinely considered the most important pests of potatoes. Potato aphid, green peach aphid, and more recently, other aphids such as the bird cherry-oat aphid Rhopalosiphum padi L. have been identified as vectors of multiple plant pathogenic viruses in potatoes. Since 2006, an area-wide trapping network consisting of ∼60 sites was developed through collaboration between researchers, extension faculty, and stakeholders, to monitor aphid populations in the Columbia Basin of Oregon (Umatilla and Morrow counties) and in northeastern Oregon (Union and Baker counties). Over a 9-yr period (2006 to 2014), aphid specimens were collected weekly using yellow bucket traps and specimens were then identified and counted to determine population levels during the growing season (May-September). Thus, aphid population data were compiled and subjected to spatial and temporal distribution analysis. Weather data, obtained from an established network of weather stations located in the monitoring areas, were used in a nonparametric multiplicative regression analysis to determine which abiotic variables may impact aphid populations. Weather conditions were characterized using confidence intervals (CIs) established based on weather data from 1999 to 2005 for each environmental variable. Aphid populations were found to have a heterogeneous distribution in most years; a few sites had high aphid populations while low numbers were observed at most sites; aphids were also found to correlate with several abiotic variables, namely, elevation, previous season temperature, and previous season dew point.

Identification of Environmental Factors Related to Claviceps purpurea Ascospore Production in Perennial Ryegrass Seed Fields and Development of Predictive Models.

Claviceps purpurea, the causal agent of ergot of perennial ryegrass seed crops, overwinters as sclerotia in the soil and releases airborne ascospores in the spring that infect flower ovaries and replace seed with sclerotia. Burkard spore traps were used to quantify the dispersal phenology and concentration of ascospores in perennial ryegrass seed fields in the Columbia Basin of Oregon. Weather factors were measured concurrently with spore trapping. Nonparametric regression, box-and-whisker plots, and univariate analysis were used to visualize and identify trends between ascospore concentrations and weather variables. Most ascospores (75.4%) were trapped when minimum soil temperatures were between 16.2 and 20.4°C. Over 67% of the total ascospores trapped were observed when minimum air temperatures were between 6.8 and 12.4°C and 64% of ascospores were trapped when daily mean dew point was between 3.7 and 8.2°C. Environmental favorability index (EFI) models were developed and validated based on their ability to predict ascospore occurrence. The EFI models were able to predict ascospore occurrence with an accuracy of 71.7 to 87.5% depending on the year. The models were up to 79.8% accurate when validated using three years of historical spore trap data not used in the EFI model development. Ninety-four percent of ascospores were trapped when cumulative air degree days, using lower and upper thresholds of 10 and 25°C, respectively, were between 230 and 403. These results suggest that weather parameters can be used to model C. purpurea ascospore occurrence and potentially improve the timing and efficacy of fungicide applications by identifying when plant protection is most needed.

Spatial Patterns of Ergot and Quantification of Sclerotia in Perennial Ryegrass Seed Fields in Eastern Oregon.

Ergot, caused by Claviceps purpurea, is a major disease of perennial ryegrass grown for seed in eastern Oregon. The objective of this research was to quantify and describe the spatial patterns of ergot severity in each of three 50-ha commercial fields of perennial ryegrass grown for seed in 2012 and 2013. In total, 1,433 and 1,405 quadrats were sampled among the three fields in 2012 and 2013, respectively, and the percentage of quadrats with ergot ranged from 59 to 90%. The mean incidence of infected seed heads in each quadrat ranged between 13 and 29%, while mean severity in each quadrat ranged from 0.2 to 0.5 sclerotia per seed head. Significant autocorrelation and clustering were observed in all three fields in both years, as indicated by Moran's I and spatial analysis by distance indices of aggregation. The mean number of ergot sclerotia collected from each field after harvest ranged between 4 and 15 sclerotia m-2 in 2012 and 18 and 119 sclerotia m-2 in 2013. Sclerotia left in perennial fields after harvest are a significant source of inoculum that should be targeted for control. This is the first study to quantify spatial patterns of ergot in perennial ryegrass and provides insights into possible mechanisms that contribute to ergot etiology and epidemiology.

Afternoon Ascospore Release in Claviceps purpurea Optimizes Perennial Ryegrass Infection.

In Kentucky bluegrass (Poa pratensis), Claviceps purpurea, the causal agent of ergot, typically releases ascospores during the early-morning hours, between about midnight and 10:00 a.m., corresponding to time of flowering, when the unfertilized ovaries are most susceptible to infection. During aeromycology studies of C. purpurea in perennial ryegrass (Lolium perenne) in northeastern Oregon during 2008 to 2010 and 2013, a strain of C. purpurea was found that released ascospores in the afternoon, coinciding with flowering in perennial ryegrass. Under controlled environmental conditions, sclerotia from perennial ryegrass and Kentucky bluegrass released spores in the afternoon and morning, respectively, consistent with timing of spore release under field conditions. Internal transcribed spacer (ITS) sequences of single sclerotial isolates from Kentucky bluegrass and perennial ryegrass were consistent with C. purpurea, although minor variations in ITS sequences among isolates were noted. Differences between Kentucky bluegrass and perennial ryegrass isolates were observed in random amplified polymorphic DNA. Evidence is provided for adaptation of C. purpurea to perennial ryegrass by means of delayed spore release that coincides with afternoon flowering in perennial ryegrass.